Method of production of adjuvant for viral vaccines

FIELD: biotechnology.

SUBSTANCE: method comprises dissolving the mixture of birch bark triterpenoids in tetrahydrofuran to obtain the solution with a concentration of 5-10 g/l. Oleic acid is dissolved in an amount of 5-10% by weight of the birch bark triterpenoids. The sterilising filtration of the mixture is carried out. 25-fold excess of 0.01 M tris buffer is added, pH 9.0±0.2, when stirring. Sonication is carried out for 5-10 min. The organic solvent is removed using ultrafiltration on hollow membranes with exclusion threshold of 300 kDa at a rate of 1.0-1.2 L/min at a pressure of 0.6-0.8 atm. Cryoprotectant is added from the group of substances: mannitol, maltose, trehalose, mannose, sorbite, sucrose. The resulting concentrated mixture is frozen.

EFFECT: invention enhances the immunogenic activity of viral vaccines and provides their stability while storage.

2 cl, 2 dwg, 6 tbl, 7 ex

 

The invention relates to biotechnology and immunology, namely the production of drugs specifically stimulating antibody formation, and can be used in medicine in the design and production of high efficient viral vaccines.

Vaccination is considered in modern terms as one of the leading mass effective means of combating infections [Results and prospects of development of vaccination in the twenty-first century, I. V. feldblyum, http://www.rusmedserv.com/epidinf/m-dokl/].

At the present time to increase the effectiveness of vaccines in experimental and clinical studies have used various adjuvants of origin: mineral hydroxide or aluminum phosphate, etc.); vegetable (saponins - QuilA, QS21); microbial (kills bacteria, lipopolysaccharide and its derivatives, CpG-DNA motifs), etc.

Due to toxicity or lack of effectiveness of most adjuvants for a wide clinical use is permitted only aluminum salts and water-oil emulsion MF-59, and in some countries, the virus-like particle (VLP - virus-likeparticles) and immune-stimulating complex (ISCOM - immunostimulatingcomplex). Foreign commercial influenza vaccines are produced with only one adjuvant MF-59, such vaccines were more immunogenic vaccines for older persons, however, had increased reactogenicity.

Development of new classes of adjuvants is a promising direction of Immunobiology. In recent decades, significant changes have occurred in the technology of production of vaccine products. Vaccines that are currently undergoing clinical trials, are significantly different from traditional vaccines. First of all, this recombinant vaccine based on purified proteins. The creation of new vaccines is, in turn, cause the search for new adjuvants. The use of adjuvants to reduce the dose of antigen in the vaccine to increase the immunogenicity of weak antigens, to prevent competition antigens in combined vaccines, to increase the rate of development and duration of the immune response in vaccinated, to induce protective properties of the mucous membranes, and also to increase the strength of the immune response in children and the elderly.

Currently in the global and domestic practice are considered adjuvants in the form of various forms of liposomes, virosomes, immunostimulatory complexes, nano-emulsion (MF59, SAF, Montanide), polymeric nanospheres, virus-like particles, etc.

The most promising adjuvants, according to the researchers, is nanoparticles (NPS). One of the reasons for the use of LF as adjuvants is the known fact that they are effectively absorbed by antigen presenting cells. Thus, if LF to bind the antigen, it will be directed absorption�hereafter macrophages, which will lead to increased immune response.

A promising direction in this field are adjuvants on the basis of birch bark triterpenoids.

When creating such adjuvants are difficulties associated with the extremely low water solubility triterpenoids. Thus, the solubility of betulin, as defined, is less than 1 μg/ml. Therefore, the establishment of water-soluble form is very important, as it should lead to an increase in particle concentration and effectiveness of an adjuvant. These forms led to the creation of spherical amorphous woofers on the basis of birch bark.

Spherical amorphous nanoparticles (SANCHO) on the basis of natural pentacyclic triterpene substances - betulin (Betulinol, Betulinol, Lupandin) are the composition of biologically active compounds. This distinguishes them from all known forms of media. They are having a pronounced spectrum of biological activity (antimicrobial, antifungal, antiviral action, hepatoprotective, anti-inflammatory, anti-cancer, anti-allergic, membranostabiliziruyuschee, immunomodulatory, antioxidant), possess adjuvant properties.

Known patent of the Russian Federation 2322091 “Composition of biologically active substances and method for producing monodisperse her”, A23L 1/30, AK 36/00, publ. 20.04.2008 and patent �Russian Federation 2322998 “Media drug and diagnostics”, AC 36/185, AC 9/10, publ. 27.04.2008 on obtaining nanodisperse of birch bark extract as a carrier of drugs and diagnostic tools. The patent describes a method of producing monodisperse, namely that in the flask with the extract of birch bark in tetrahydrofuran (5 mg/ml) with vigorous stirring was added distilled water. The resulting dispersion was stirred, the solvent was evaporated on a rotary evaporator at a temperature not exceeding 40°C. Then nanodispersion was treated in an ultrasonic bath and concentrated on a rotary evaporator to 1-1.5 mg/ml. the Average size of the nanoparticles obtained amounted to 187 nm.

The disadvantage of this method is unsuitable for industrial applications is the difficulty of scaling the process, the limiting factor is the removal of the organic solvent on a rotary evaporator.

Known patent of the Russian Federation 2424516 “a Method of separating the mixture to obtain an aqueous dispersion of spherical nanoparticles”, G01N 33/15, WV 1/00, publ. 20.07.2011 by selection of the mixture to obtain an aqueous dispersion of spherical nanoparticles from a mixture of poorly soluble in water the birch bark triterpenoids, including injection of excess water in the mortar birch bark triterpenoids in water-miscible organic solvent (tetrahydrofuran) with formation of a dispersion containing spherical nanoparticles and crystals triterpenoids from birch bark, The resulting dispersion is centrifuged, separating the crystals from the fraction of spherical nanoparticles separated nanoparticles is evaporated to obtain a solid mixture of triterpenoids for the formation of morphologically homogeneous spherical nanoparticles by re-injection.

The disadvantage of this method is the lack of purification of tetrahydrofuran. According to the classification of organic solvents tetrahydrofuran belongs to the second class phenoloxidase solvents and its contents in the drugs need to be monitored.

Known patent of the Russian Federation 2355423 “Adjuvant”, A61K 47/06, C08H 5/04, publ. 20.05.2009, which discloses a method of preparation of adjuvants on the basis of betulin nanoparticles. According to a known method to obtain adjuvant in a 0.5% aqueous solution of betulin nanoparticles in the form of dried drug nanoparticles extract of birch bark. Adjuvants prepared as follows. Dissolve birch bark dry extract (BAS-65) in an organic solvent at a concentration of 2.5-5 g/l; adding to the said solution a large quantity of water (up to 25 volumes of water relative to the volume of the solution); the solvent is removed and the bulk of the water. To the resulting aqueous solution of the nanoparticles add the cryoprotectant (e.g., sorbitol), and perform lyophilization. Before �Manizales adjuvants dissolved in phosphate buffer (pH 7,5), treated with ultrasound for 15 minutes, then cooled.

The disadvantage is the complexity of the scaling process, the limiting factor is the removal of the organic solvent on a rotary evaporator.

As the closest analogue may be specified a method of producing an adjuvant for vaccines in the form of spherical amorphous nanoparticles (SANCHO) from a mixture of birch bark triterpenoids (Gavrilov L. A., NANOPARTICLES of HYDROPHOBIC NATURAL COMPOUNDS AS ADJUVANTS, dissertation, Moscow, 2011). Adjuvant obtained by depositing the excess water from water-miscible solvents, such as tetrahydrofuran, adding Miramistin or oleic acid to 2% by weight triterpenoids, removing the organic solvent by rotary evaporator, the lyophilization in the presence of cryoprotectant. Additionally, the evaporation may be used ultrafiltration. The sizes of spherical particles are 100-400 nm. However, although this method enables to increase the stability and effectiveness of an adjuvant, in its practice, there are losses of triterpenoids.

In addition, in the described method, there is no stage obtaining sterilization, which is one of the main requirements for drugs administered parenterally. In accordance with the European Pharmacopoeia and with the applicable R�Russia regulatory documents preparations for parenteral administration must be sterile non-pyrogenic, non-toxic.

The features of the prototype, coinciding with the features of the invention consist in the presence of a dissolved mixture of birch bark triterpenoids in tetrahydrofuran, adding oleic acid, removal of tetrahydrofuran, the addition of cryoprotectant and lyophilization.

The problem solved by the invention is to improve the quality of an adjuvant, allowing to receive it in accordance with the requirements of the drugs administered parenterally: sterility, pyrogen-free, toksichnosti.

The technical result, provides solution to the aforementioned problem, is the removal of the toxicity of the drug, reducing the residual concentration of THF, to ensure its sterility, apirogennost, scalability and stability during storage.

The technical result is achieved in that in the method of obtaining adjuvant comprising dissolving a mixture of birch bark triterpenoids in tetrahydrofuran, the addition of oleic acid, removal of tetrahydrofuran, the addition of cryoprotectant and freeze drying according to the invention a mixture of birch bark triterpenoids in tetrahydrofuran with a concentration of 5-10 g/l followed by dissolving oleic acid in an amount of 5-10% by weight of birch bark triterpenoids, spend sterilizing filtration of the mixture, form a homogeneous dispersion of spherical� amorphous nanoparticles by adding a 25-fold excess of 0.01 M Tris-buffer, pH 9,0±0,2, with stirring, followed by ultrasonic treatment for 5-10 min, remove the tetrahydrofuran by using ultrafiltration hollow membranes with the exception threshold of 300 kDa at a speed of 1.0-1.2 l/min, at a pressure of 0.6-0.8 ATM add the cryoprotectant from the group of substances: mannitol, maltose, trehalose, mannose, sorbitol, sucrose, frozen obtained concentrated mixture containing a mixture of terpenoids 1 mg/ml temperature below -35°C, maintained at this temperature for 4-6 hours and lyophilizers, the lyophilization is carried out at a temperature of -35°C for 15 hours; followed by drying at 20-25°C for 15 hours.

Preferably in the method for lyophilization to use sucrose as the cheapest carbohydrate.

The inclusion of the stage of processing the ultrasound helped to a greater extent adjuvant to stabilize.

Important to practice mode lyophilization is the determination of eutectic temperature at which the aqueous solution is transferred to the solid phase. The importance of this stage is determined by the fact that the presence of a liquid phase, even in a seemingly frozen material in the early phase sublimation can lead to irreversible phenomena, such as the violation of the integrity of the layer of the drug and its physical structure. The eutectic point for a spherical amorphous nanoparticles was determined from the analysis of the quantity�and electrical resistance on temperature. The lyophilization of the samples was carried out on the machine for sublimirovanny TG-50 (Germany). According to the analysis of definitions of the eutectic point, it was found that spherical amorphous nanoparticles with a concentration of cryoprotectant 1/10-1/20 by weight have a eutectic temperature of -15±6°C. optimally carry out the stage of lyophilization at a temperature of drug -35°C, the duration of the stage 15 hours; the dried adjuvant drug in freezing temperatures 20-25°C for 15 hours.

The presence of the adjuvant composition of oleic acid in an amount of 5-10% by weight of birch bark triterpenoids, ensures stability during storage, the introduction of the stage sterile filtration provided the sterile and non-pyrogenic adjuvant, the use of ultrafiltration purification was facilitated by the exclusion of toxicity and removal of tetrahydrofuran, lower costs. The set of distinctive features provides easy scaling of the technological process from laboratory to industrial.

New features of the invention are that a mixture of birch bark triterpenoids in tetrahydrofuran with a concentration of 5-10 g/l, followed by dissolving oleic acid in an amount of 5-10% by weight of birch bark triterpenoids, spend sterilizing filtration of the mixture, form a homogeneous dispersion of spherical �polymorphic nanoparticles by adding a 25-fold excess of 0.01 M Tris-buffer, pH 9,0±0,2, with stirring, followed by ultrasonic treatment for 5-10 min, remove the tetrahydrofuran by using ultrafiltration hollow membranes with the exception threshold of 300 kDa at a speed of 1.0-1.2 l/min, at a pressure of 0.6-0.8 ATM, add the cryoprotectant from the group of substances: mannitol, maltose, trehalose, mannose, sorbitol, sucrose, frozen obtained concentrated mixture containing a mixture of terpenoids 1 mg/ml temperature below -35°C, maintained at this temperature for 4-6 hours and lyophilizers, the lyophilization is carried out at a temperature of -35°C for 15 hours, followed by drying at 20-25°C for 15 hours.

The invention is illustrated by drawings, where Fig. 1 shows the freezing of spherical amorphous nanoparticles; Fig. 2 - mode sublimation spherical amorphous nanoparticles.

The invention can be demonstrated by the following examples below.

Example 1. Receiving adjuvant viral vaccines

For the purpose of obtaining adjuvant that meets the requirements for preparations for parenteral administration, we worked out the technology in which all initial solutions must be sterilized at the initial stages of manufacture of an adjuvant, and the entire future of the technological process should take place under sterile conditions. Therefore, the first stage is carried out Rast�the working mixture of birch bark triterpenoids, containing, wt.%: betulin 65-71, lupeol 12-16, 3-O-caveat betulin 5-15 in tetrahydrofuran. To this mixture was added oleic acid in the amount of 5-10% by weight of birch bark triterpenoids. Sterilized mixture obtained using the modified filter element surface charge, are resistant to organic solvents, such as a nylon membrane with a pore diameter of 0.22 μm brand NRG Pall N66+c.

With the aim of obtaining a homogeneous dispersion of spherical amorphous nanoparticles mixture is added to sterile 25-fold volume of sterile buffer (pH 9,0±0,2) using a peristaltic pump with constant stirring for 15 minutes overhead stirrer propeller type. The subsequent homogenization by ultrasound for 5 min was formed spherical amorphous nanoparticles. Thus observed the destruction of the aggregated particles, pop up to the surface, and the variance was homogeneous.

For the purpose of removal of the organic solvent used ultrafiltration on hollow fibers with a nominal cut-off molecular weight of 300 kDa. Purification from organic solvents is optimized by the use of 0.01 M Tris-buffer solution with a pH of 9.0±0.2 for 2-3-fold dilution of the initial dispersion of spherical amorphous nanoparticles. The rate of ultrafiltration composition�Yala 1,0-1,2 l/min, the pressure set in the range of 0.6 to 0.8 ATM. Nanodispersions concentrated to a content of the basic substance of 1 mg/ml.

Lyophilization: freeze the drug is below the temperature of -35°C and maintained at this temperature for at least 4-6 hours; conducting stage lyophilization at a temperature of drug -35°C, the duration of the stage 15 hours; the dried drug in freezing temperatures 20-25°C for 15 hours (Fig.1, 2).

Appearance of the obtained freeze-dried adjuvant was a porous mass of white or yellowish-white, with a residual humidity (3,5±0,5) %.

Residual THF in the finished product is determined by GLC. Analysis of the results showed that the used ultrafiltration method provides effective removal of THF, the concentration in the preparation was 0,176±0,009 mg/ml. Subsequent stage of the lyophilization contributed to a decrease in the residual concentration of THF to 0,122±0.004 mg/ml, i.e. almost 30%. It should be noted that according to the global Fund XII edition, the maximum allowable amount of tetrahydrofuran taken in the composition of a daily dose of medicinal products, should not exceed 7.2 mg/ml in 2360 times less than when administered 25 µg, SANCHO (the intended content per dose of vaccine preparation).

The proposed method of obtaining spherical amorphous nanoparticles, having�x Zeta potential disadvantage of 44.3 mV and the average size of 160-180 nm, provides the conditions under which it is not formed crystalline nanoparticles.

Example 2. The resuspension of lyophilized dried adjuvant drugs

Freeze-dried preparation was resuspended in water for injections. The resulting solution was stirred for 2-3 min, for example, using a shaker or stirrer, avoiding churning. Also recommended treatment of the suspension by ultrasound for 3-5 minutes.

Example 3. Test for pyrogenicity and sterility

To test for pyrogenicity rehydrated adjuvant was administered to the rabbits at 50 mg / kg intravenously. The maximum amount of temperature change at three rabbits was 0.9°C (tab.1), which shows that apirogennost adjuvant.

Table 1
The results of the production test for pyrogenicity adjuvant
No. rabbitThe weight of the rabbit, kgTemperature prior to the introduction, °CThe temperature in an hour, °CThe temperature after 2 hours, °CThe temperature after 3 hours, °CThe maximum amount of temperature change, °C
12,75of 39.439,339,539,50,9
22,6539,338,939,039,2
32,5039,539,139,239,3

Definition of sterility adjuvant was carried out in accordance with GF XII edition with two modes of 22.5±2.5°C and 32.5±2,5°C. it Was found that adjuvant sterile.

Thus, the developed technology allows to obtain sterile and non-pyrogenic adjuvant that meets the requirements for preparations for parenteral administration and allows their use as vaccine adjuvants.

Example 4. Evaluation of sorption ability of an adjuvant for example diphtheria, tetanus toxoids and surface antigen of hepatitis b virus

One of the main mechanisms of action of adjuvants is a "depot" effect, which is implemented by the binding of antigen and adjuvant. The same sorption ability of an adjuvant can�t be through electrostatic interactions.

When mining technologies was determined sorption capacity adjuvant for example diphtheria, tetanus toxoids, surface antigen of hepatitis b virus. To obtain the vaccine compositions selected the necessary amount of adjuvant with a pH of 9.00 and was added a calculated amount of the antigen was mixed and left for 30 min for adsorption. Then brought in 0.9% sodium chloride solution to obtain vaccine dose. Prepared samples of vaccines centrifuged at 10,000 rpm, supernatant was separated, which was analyzed by ELISA and the results of the titration was determined by the sorption capacity of an adjuvant (table.2).

Table 2
Determination of sorption capacity adjuvant
The adjuvant concentration, mcg/mlThe efficiency of incorporation of antigens, %
diphtheria toxoidtetanus toxoidHBsAg
503,595,8496,56
1001,24,34 97,85
2009,913,1198,92
40013.56 MHzat 14.7399,47
50029,3520,9899,39

As can be seen from table 2, an adjuvant in a concentration of 50 μg/ml effectively binds a surface antigen of the virus of hepatitis B. In turn, even adjuvant at a concentration of 500 μg/ml, the efficiency of binding to diphtheria and the tetanus toxoid is less than 30%.

Example 5. The study of the adjuvant properties on the example of the vaccine against hepatitis B

Adjuvant properties were studied on the model of the vaccine against hepatitis B in experiments on Guinea pigs weighing 350-400 g. subcutaneously twice with an interval of 14 days was administered the hepatitis B vaccine with different adjuvants in a volume of 0.5 ml (10 mcg HBsAg). On the 14th and 28th day after immunization in sera of Guinea pigs by enzyme immunoassay was determined antibodies to surface antigen of the hepatitis b virus.

Table 3
The results of the study of immunogenic properties of the vaccine against hepatitis B with adjuvants of different nature in experiments on Guinea pigs
The vaccine composition (total dose per animal)Geometric mean titer (SGT), mIU/ml
I immunizationII immunization
20 µg HBsAg + 50 μg adjuvant40,02*1276,63**
[17,53-91,35][830,24-1963,03]
20 µg HBsAg + 500 mcg adjuvant44,92*1192,26**
[15,40-130,96][681,91-2084,57]
Control of a commercial hepatitis B vaccine (20 µg HBsAg + 500 mcg Al(OH)3)22,81586,69
[10,94-47,52][401,97-856,30]
* Differences of mean values is not significant compared with control; ** Differences of mean values is significant in comparison with the control.

In our studies, it was found that the experimental�experimental adjuvant at concentrations much lower, it is widely used adjuvant - gel aluminum hydroxide, provides a high immune response to the vaccine against hepatitis B.

Example 6. The study of the adjuvant properties on the example of a vaccine against influenza virus

In the experiment with pandemic influenza vaccines used white mice with body weight of 10-13 g. Immunization was performed as follows: in each of the hind paws of mice was injected with 100 µl of freshly prepared solutions (with or without adjuvants) by intramuscular injection of 1.5 μg of hemagglutinin (HA) on the animal. Blood sampling was performed one month after immunization. The specific activity in sera of mice were determined by ELISA.

Table 4
Immunogenicity of influenza vaccine with an experimental adjuvant
Vaccine compositionsThe amount of hemagglutinin (HA) per animal, ugThe geometric mean of the absorbance value
1.5 μg HA + 25 μg adjuvant1,51,015[0,877-1,173]**
1.5 μg HA + 250 µg adjuvant1,5 0,791[0,596-1,051]**
1.5 μg HA + 500 mcg adjuvant1,50,676[0,546-0,836]***
1.5 µg HA*1,50,551[0,429-0,708]
* Control (rezorbirovanny hemagglutinin); ** Differences of mean values is significant in comparison with control; *** Differences of mean values is not significant compared to control.

Presents the results of ELISA showed that the experimental adjuvant showed pronounced adjuvanting properties. The maximum level of antibodies in animals has provided a vaccine composition with a low dose of an experimental adjuvant.

Example 7. The study of the toxic properties of the experimental adjuvant

Toxic properties of the drug was evaluated in experiments on determination of acute and chronic toxicity. To study acute toxicity to white mice were injected intraperitoneally a single dose of 0.5 ml (25 µg/dose) mouse (which corresponds 2994 human doses), Guinea pigs (weighing 300-350 g) subcutaneously at a dose of 5.0 ml (250 mcg/dose) on a Guinea pig (which corresponds to 1851 the human dose). In the study of chronic toxicity to white mice were injected adjuvant at a concentration of 50 μg/ml at a dose of 0.5 ml (25 µg) VNU�rebrushing triple (1, 4, day 9) in a total dose of 1.5 ml per mouse (corresponding 8982 human dose). Animals of group drove a 0.9% solution of sodium chloride by similar schemes.

The research findings showed that single and repeated administration of the drug did not cause the death of animals, resulted in decrease in body weight (table.5, 6), changes in hair distribution, necrobiotic changes at the injection site.

367,00±3,50*
Table 5
Dynamics of changes in the weight of the animals after a single injection of the drug
The study drugType of animalThe initial mass of animals, gThe weight of the animals at 1 d, gThe weight of the animals at 8 days, g
Experimental adjuvantWhite mouse18,24±0,1418,58±0,184*21,48±1,24*
Guinea pigs317,50±2,10320,50±1,90*
NaCl 0,9% (control)White mouseto 18.09±0,1618,30±0,1422,20±0,98
Guinea pigs324,10±2,39327,50±2,03372,10±4,15
* p>0.05 insignificant difference compared to control.
Table 6
Dynamics of change in the mass of white mice after repeated administration
The study drugThe initial mass of animals, gThe weight of the animals at 1 d, gThe weight of the animals at 22 d, g
Experimental adjuvant18,46±0,1818,78±0,60*27,80±2,89*
NaCl 0,9% (control)18,60±0,46 carries 18.68±0,6227,63±2,80
* p>0.05 insignificant difference compared to control.

After 24 h, and 13 days after the last injection in the evaluation of chronic toxicity in the comparison group and experimental group, the internal organs of animals had all the characteristics, location and structure. Membranes lining the internal cavity moist, grayish-pink color, without signs of inflammation.

Thus, in the study of acute and chronic toxicity, it was found that experimental adjuvant does not cause symptoms of intoxication, does not have a toxic effect on the systems and organs of laboratory animals, is not conducive to the development of pathological, including inflammatory, degenerative and necrotic changes.

1. A method of producing an adjuvant for vaccines comprising dissolving a mixture of birch bark triterpenoids in tetrahydrofuran, the addition of oleic acid, removal of tetrahydrofuran, the addition of cryoprotectant and lyophilization, characterized in that a mixture of birch bark triterpenoids in tetrahydrofuran with a concentration of 5-10 g/l, followed by dissolving oleic acid in an amount of 5-10% by weight of birch bark triterpenoids, spend sterilizing filtration of the mixture, form a homogeneous dispersion of spherical �polymorphic nanoparticles by adding a 25-fold excess of 0.01 M Tris-buffer, pH 9,0±0,2, with stirring, followed by ultrasonic treatment for 5-10 min, remove the tetrahydrofuran by using ultrafiltration hollow membranes with the exception threshold of 300 kDa at a speed of 1.0-1.2 l/min, at a pressure of 0.6-0.8 ATM, add the cryoprotectant from the group of substances: mannitol, maltose, trehalose, mannose, sorbitol, sucrose, frozen obtained concentrated mixture containing a mixture of terpenoids 1 mg/ml temperature below -35°C, maintained at this temperature for 4-6 hours and lyophilizers, the lyophilization is carried out at a temperature of -35°C for 15 hours followed by drying at 20-25°C for 15 hours.

2. A method according to claim 1, characterized in that the use of cryoprotectant sucrose.



 

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FIELD: medicine, pharmaceutics.

SUBSTANCE: group of inventions refers to oral care compositions containing a basic amino acid or its salt. The presented oral care composition promoting the dentine defect closure in the oral cavity contains arginine in the free form or in the form of a salt, and an abrasive substance representing synthetic amorphous silica and containing small particle fractures making at least approximately 5% of total weight of the composition, wherein the particles of the small particle fracture having d50 from 3 to 4 mcm. What is also presented is a method of treating sensitive teeth in the oral cavity involving using the oral treatment with this composition, as well as using arginine as a part of the oral care composition and for producing a therapeutic agent, wherein the above composition and agent contain the above abrasive material containing the small particle fracture.

EFFECT: group of inventions provide the effective dentin defect closure in the patient's oral cavity.

8 cl, 1 tbl, 6 ex

FIELD: medicine, oncology, amino acids.

SUBSTANCE: invention relates, in particular, to the development of an antitumor preparation based on natural substances. Invention relates to an amino acid preparation comprising at least one modified essential amino acid obtained by treatment of amino acid by ultraviolet radiation (UV) at wavelength 250-350 nm for 12-80 h at temperature 15-30oC or with ozone at temperature 15-25oC. The modified amino acid has no toxicity for health cells. Also, invention relates to a method for preparing such preparation. Invention provides the development of an antitumor preparation based on modified amino acids and expanded assortment of antitumor preparations being without cytotoxicity for normal cells.

EFFECT: valuable medicinal antitumor properties of preparation.

8 cl, 4 tbl, 2 dwg, 4 ex

FIELD: medicine, gastroenterology.

SUBSTANCE: after selecting all biliary portions one should introduce firstly a certain anolyte at the quantity of 100-150 ml at ORP = +700 - +1200 mV; pH = 6.0 - 7.3 for a patient through a sound, then one should introduce a certain catholyte at the quantity of 150-200 ml at ORP being -400 - -800 mV; pH = 8.5 - 9.5 into a free sound. The present innovation enables to decrease relapses in diseases of biliary canals at the background of restricted usage of chemotherapeutical preparations.

EFFECT: higher efficiency of curative sounding.

2 ex, 1 tbl

FIELD: organometal chemistry, stomatology.

SUBSTANCE: invention relates to titanium compounds of formula [TiF2L2] or containing group of formula [TiFxLy]z-, wherein x = 2, 4 or 5; y = 1 or 2; z = 1 or 2; L is group of formula II , wherein m = 1; n = 0, 1 or 2, in enantiomer, diastereomer forms, or mixture thereof, including racemates; as well as free bases or pharmaceutically acceptable salts. Composition containing claimed compounds for buccal application to prevent dental caries also is disclosed.

EFFECT: compounds with improved stability capable to form lacquer on dental surface.

14 cl, 1 ex

FIELD: cosmetology, food industry.

SUBSTANCE: the present innovation deals with compositions for cosmetic preparations, means for personal hygiene, purifying components, biocidal components, functional food and food additives. Due to these compositions one should introduce bioactive glass containing SiO2, CaO, P2O5 and Na2O, not obligatory, into the objects mentioned. Moreover, the present innovation deals with methods to obtain and apply the compositions mentioned. It enables to obtain cosmetic products capable to withstand bacterial infection.

EFFECT: higher efficiency.

134 cl, 19 ex, 5 tbl

FIELD: medical engineering.

SUBSTANCE: method involves acting in turn with ultrasonic and magnetic field with switch frequency of 10 to 160 Hz. Ultrasonic field intensity is set below cavitation threshold and ultrasonic oscillation frequency being equal to liquid acoustic relaxation frequency. Magnetic field induction being selected within the limits of 30-50 mTesla units.

EFFECT: enhanced effectiveness of liquid activation; reduced power consumption.

1 dwg

FIELD: medicine, infectious diseases.

SUBSTANCE: method involves administration of electroactivated aqueous solutions of salts with negative and positive redox-potential and definite pH value and with the concentration of active chlorine in the definite sequence and regimen. Invention promotes to arresting basic pathogenetic syndromes of disease due to normalization of metabolic processes in body and recovery of the normal microbiocenosis and by development of conditions promoting to using the own natural potential by human. Invention can be used for complex treatment of parasitic and infectious diseases.

EFFECT: improved method for treatment.

3 cl, 4 ex

FIELD: experimental medicine, immunology.

SUBSTANCE: invention relates to a method for immunization of intact animals with ram erythrocytes treated by alternating magnetic field 50 Hz and 25 mTl for 30 min. Method provides enhancement of induction of the primary immune response in experimental animals. Invention can be used for stimulation of immune response in experiment.

EFFECT: improved method for stimulating.

2 tbl

FIELD: medicine, stomatology.

SUBSTANCE: invention proposes an agent for prophylaxis and treatment of dental caries that comprises carrier and active substance wherein carrier is made as a bicomponent system consisting of rubber or rubber-like elastomer eliciting high residual deformation and spherical or approaching to spherical rubber particles or rubber-like polymer eliciting the residual deformation less 0.001% and showing deformation indices 10-25 kg/cm2 at relative elongation 0.1-0.25 and in the ratio of components in the carrier = (30:70)-(85:15) wt.-%, size of particles is 1.0-5.0 mm and wherein active substance involves active sources of fluorine, calcium and phosphate ions. Agent is made as plate, rod or block. Method for prophylaxis and treatment of caries involves chewing the indicated agent with strength providing release of active substance from carrier and maintaining the concentration of calcium and fluorine ions in saliva 10-4 - 10-5 M, respectively, for 5-15 min, not less. Agent and method show high effectiveness in treatment and prophylaxis of caries and comfortable using also.

EFFECT: improved, enhanced and valuable medicinal properties of agent and method.

3 cl, 2 tbl

FIELD: medicinal biotechnology, immunology.

SUBSTANCE: invention relates to producing medicinal biological preparations. Invention relates to a method for enhancing specific activity of immunoglobulins of class G. Method involves freezing immunoglobulin G-containing fractions to temperature from -20°C to -196°C and subjected for gamma-irradiation in doses 5-50 kGr with power of absorbed dose 60-110 Gr/min. method provides elevating the specific activity of immunoglobulin G, increasing titer, avidity and reducing anti-complementary activity.

EFFECT: improved method for elevating activity of antibodies.

3 tbl, 3 dwg, 4 ex

FIELD: medicine, stomatology.

SUBSTANCE: therapy suggested should be performed due to applying a thin layer of 0.1%-butyric estradiol solution followed by a 5-min-long ultrasound treatment of the target surface. The method provides complex therapy and prophylaxis of parodontium diseases due to local substitution hormonotherapy with estradiol-containing solution.

EFFECT: higher efficiency of therapy.

1 ex

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